FREE SHIPPING over $40
OVER 10000 FIVE-STAR REVIEWS
5% OFF ALL SUBSCRIPTIONS
100% HAPPINESS GUARANTEE
Analyze Diet
0
Journal of veterinary internal medicine2021; 35(2); 1131-1139; doi: 10.1111/jvim.16065

Fibrinogen heterogeneity in horses.

Abstract: Fibrinogen heterogeneity has been observed in humans and can influence fibrinogen measurements when using the modified Clauss assay. We hypothesized that fibrinogen heterogeneity also exists in horses. Objective: To determine whether fibrinogen heterogeneity exists in horses. Methods: Five clinically healthy horses from the university equine teaching herd. Methods: Presumed fibrinogen was purified from pooled citrated plasma and electrophoresis performed. The purified protein was subjected to Western blotting using sheep antiserum against human fibrinogen, and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Results: Gel electrophoresis of nonreduced equine purified protein yielded 2 protein bands (approximately 377 and 318 kDa) that corresponded with the molecular weights of human high molecular weight fibrinogen and low molecular weight fibrinogen fractions, respectively. Electrophoretograms of reduced purified protein, Western blots, and LC-MS/MS supported that the purified nonreduced protein bands were fibrinogen. Conclusions: Fibrinogen heterogeneity exists in horses.
© 2021 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals LLC. on behalf of the American College of Veterinary Internal Medicine.
Get Full Text
Publication Date: 2021-02-19 PubMed ID: 33604912PubMed Central: PMC7995376DOI: 10.1111/jvim.16065Google Scholar: Lookup
The Equine Research Bank provides access to a large database of publicly available scientific literature. Inclusion in the Research Bank does not imply endorsement of study methods or findings by Mad Barn.
LinkedInCopy
  • Journal Article

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

The researchers in this study identified different forms or variants (heterogeneity) of the protein fibrinogen in horses, much like what has been observed in humans. Their findings could influence how fibrinogen levels are measured in horses.

Research Objective

The main aim of this research was to establish whether there are different forms of fibrinogen (a blood-clotting protein) in horses, in a similar fashion to humans. This phenomenon is referred to as fibrinogen heterogeneity. Finding out that fibrinogen has different forms in horses could possibly influence how its levels are measured.

Research Methodology

  • Five clinically healthy horses were sourced from the university’s equine teaching herd for blood withdrawal.
  • The blood drawn was treated with an anticoagulant (citrated), and fibrinogen was extracted from the plasma – the liquid part of the blood.
  • The purified fibrinogen was then subjected to gel electrophoresis – a lab technique used to separate proteins based on their size and charge. This was aimed at identifying potential fibrinogen variants.
  • The proteins identified through gel electrophoresis were further analysed using Western blotting – a technique to detect specific proteins – and using an antiserum against human fibrinogen.
  • They also used liquid chromatography-tandem mass spectrometry (LC-MS/MS) to perform a detailed analysis of the proteins.

Research Findings

  • When the researchers ran the purified protein samples through gel electrophoresis, they found two protein bands, which suggest two different forms or variants of fibrinogen in horses.
  • The weights of the protein bands corresponded with the weights of different forms of fibrinogen observed in humans — high molecular weight and low molecular weight fibrinogen.
  • The results from Western blot and LC-MS/MS analyses further confirmed that the protein bands were indeed variants of fibrinogen.

Conclusion

The researchers concluded that fibrinogen heterogeneity exists in horses, consistent with their research hypothesis. This adds to the understanding of the protein composition of horse blood and may influence how fibrinogen levels are measured.

Cite This Article

APA
Russell EB, Courtman NF, Santos LL, Tennent-Brown BS. (2021). Fibrinogen heterogeneity in horses. J Vet Intern Med, 35(2), 1131-1139. https://doi.org/10.1111/jvim.16065

Publication

Journal of veterinary internal medicine
ISSN: 1939-1676
NlmUniqueID: 8708660
Country: United States
Language: English
Volume: 35
Issue: 2
Pages: 1131-1139

Researcher Affiliations

Russell, Elise B
  • U-Vet Werribee Animal Hospital and Faculty of Veterinary and Agricultural Sciences, The University of Melbourne, Werribee, Victoria, Australia.
Courtman, Natalie F
  • U-Vet Werribee Animal Hospital and Faculty of Veterinary and Agricultural Sciences, The University of Melbourne, Werribee, Victoria, Australia.
Santos, Leilani L
  • U-Vet Werribee Animal Hospital and Faculty of Veterinary and Agricultural Sciences, The University of Melbourne, Werribee, Victoria, Australia.
Tennent-Brown, Brett S
  • U-Vet Werribee Animal Hospital and Faculty of Veterinary and Agricultural Sciences, The University of Melbourne, Werribee, Victoria, Australia.

MeSH Terms

  • Animals
  • Blotting, Western / veterinary
  • Chromatography, Liquid / veterinary
  • Fibrinogen
  • Horses
  • Sheep
  • Tandem Mass Spectrometry / veterinary

Grant Funding

  • University of Melbourne

Conflict of Interest Statement

Authors declare no conflict of interest.

References

This article includes 31 references
  1. Weisel JW, Dempfle CH. Fibrinogen structure and function. Hemostasis and Thrombosis: Basic Principles and Clinical Practice 6th ed. Philadelphia, PA: Lippincott WIlliams & WIlkins; 2012:254‐271.
  2. Holm B, Godal HC. Quantitation of the three normally-occurring plasma fibrinogens in health and during so-called "acute phase" by SDS electrophoresis of fibrin obtained from EDTA-plasma.. Thromb Res 1984 Aug 1;35(3):279-90.
    pubmed: 6431629doi: 10.1016/0049-3848(84)90359-1google scholar: lookup
  3. Holm B, Nilsen DW, Kierulf P, Godal HC. Purification and characterization of 3 fibrinogens with different molecular weights obtained from normal human plasma.. Thromb Res 1985 Jan 1;37(1):165-76.
    pubmed: 3983897doi: 10.1016/0049-3848(85)90043-xgoogle scholar: lookup
  4. Adler G, Duchinski T, Jasinska A, Piotrowska U. Fibrinogen fractions in the third trimester of pregnancy and in puerperium.. Thromb Res 2000 Mar 15;97(6):405-10.
    pubmed: 10704649doi: 10.1016/s0049-3848(99)00190-5google scholar: lookup
  5. Holm B, Nilsen DW, Godal HC. Evidence that low molecular fibrinogen (LMW) is formed in man by degradation of high molecular weight fibrinogen (HMW).. Thromb Res 1986 Mar 15;41(6):879-84.
    pubmed: 3705020doi: 10.1016/0049-3848(86)90387-7google scholar: lookup
  6. Nakashima A, Sasaki S, Miyazaki K, Miyata T, Iwanaga S. Human fibrinogen heterogeneity: the COOH-terminal residues of defective A alpha chains of fibrinogen II.. Blood Coagul Fibrinolysis 1992 Aug;3(4):361-70.
    pubmed: 1420813
  7. Jensen T, Halvorsen S, Godal HC, Sandset PM, Skjøonsberg OH. Discrepancy between fibrinogen concentrations determined by clotting rate and clottability assays during the acute-phase reaction.. Thromb Res 2000 Dec 1;100(5):397-403.
    pubmed: 11150581doi: 10.1016/s0049-3848(00)00344-3google scholar: lookup
  8. Lipinska I, Lipinski B, Gurewich V, Hoffmann KD. Fibrinogen heterogeneity in cancer, in occlusive vascular disease, and after surgical procedures.. Am J Clin Pathol 1976 Dec;66(6):958-66.
    pubmed: 998569doi: 10.1093/ajcp/66.6.958google scholar: lookup
  9. Lipinski B, Lipinska I. Effect of magnesium on fibrin formation from lower molecular weight (LMW) fibrinogen.. Magnes Res 2000 Dec;13(4):233-7.
    pubmed: 11153893
  10. Reganon E, Vila V, Ferrando F, Martínez-Sales V, Fayos L, Ruano M, Aznar J. Elevated high molecular weight fibrinogen in plasma is predictive of coronary ischemic events after acute myocardial infarction.. Thromb Haemost 1999 Nov;82(5):1403-5.
    pubmed: 10595627
  11. CLAUSS A. [Rapid physiological coagulation method in determination of fibrinogen].. Acta Haematol 1957 Apr;17(4):237-46.
    pubmed: 13434757doi: 10.1159/000205234google scholar: lookup
  12. Millar HR, Simpson JG, Stalker AL. An evaluation of the heat precipitation method for plasma fibrinogen estimation.. J Clin Pathol 1971 Dec;24(9):827-30.
    pmc: PMC477193pubmed: 5003786doi: 10.1136/jcp.24.9.827google scholar: lookup
  13. Mackie IJ, Kitchen S, Machin SJ, Lowe GD. Guidelines on fibrinogen assays.. Br J Haematol 2003 May;121(3):396-404.
    pubmed: 12716361doi: 10.1046/j.1365-2141.2003.04256.xgoogle scholar: lookup
  14. Vila V, Regañón E, Llopis F, Aznar J. A rapid method for isolation of fibrinogen from human plasma by precipitation with polyethylene glycol 6,000.. Thromb Res 1985 Sep 1;39(5):651-6.
    pubmed: 3936222doi: 10.1016/0049-3848(85)90248-8google scholar: lookup
  15. Smith PK, Krohn RI, Hermanson GT, Mallia AK, Gartner FH, Provenzano MD, Fujimoto EK, Goeke NM, Olson BJ, Klenk DC. Measurement of protein using bicinchoninic acid.. Anal Biochem 1985 Oct;150(1):76-85.
    pubmed: 3843705doi: 10.1016/0003-2697(85)90442-7google scholar: lookup
  16. Manten GT, Sikkema JM, Franx A, Hameeteman TM, Visser GH, de Groot PG, Voorbij HA. Increased high molecular weight fibrinogen in pre-eclampsia.. Thromb Res 2003;111(3):143-7.
    pubmed: 14678811doi: 10.1016/j.thromres.2003.08.025google scholar: lookup
  17. Manten GT, Franx A, Sikkema JM, Hameeteman TM, Visser GH, de Groot PG, Voorbij HA. Fibrinogen and high molecular weight fibrinogen during and after normal pregnancy.. Thromb Res 2004;114(1):19-23.
    pubmed: 15262480doi: 10.1016/j.thromres.2004.04.008google scholar: lookup
  18. Ignjatovic V, Straka E, Summerhayes R, Monagle P. Age-specific differences in binding of heparin to plasma proteins.. J Thromb Haemost 2010 Jun;8(6):1290-4.
    pubmed: 20218985doi: 10.1111/j.1538-7836.2010.03847.xgoogle scholar: lookup
  19. Ignjatovic V, Lai C, Summerhayes R, Mathesius U, Tawfilis S, Perugini MA, Monagle P. Age-related differences in plasma proteins: how plasma proteins change from neonates to adults.. PLoS One 2011 Feb 18;6(2):e17213.
    pmc: PMC3041803pubmed: 21365000doi: 10.1371/journal.pone.0017213google scholar: lookup
  20. Goodman JK, Zampronio CG, Jones AME, Hernandez-Fernaud JR. Updates of the In-Gel Digestion Method for Protein Analysis by Mass Spectrometry.. Proteomics 2018 Dec;18(23):e1800236.
    pmc: PMC6492177pubmed: 30259661doi: 10.1002/pmic.201800236google scholar: lookup
  21. Rosenfeld J, Capdevielle J, Guillemot JC, Ferrara P. In-gel digestion of proteins for internal sequence analysis after one- or two-dimensional gel electrophoresis.. Anal Biochem 1992 May 15;203(1):173-9.
    pubmed: 1524213doi: 10.1016/0003-2697(92)90061-bgoogle scholar: lookup
  22. Shevchenko A, Tomas H, Havlis J, Olsen JV, Mann M. In-gel digestion for mass spectrometric characterization of proteins and proteomes.. Nat Protoc 2006;1(6):2856-60.
    pubmed: 17406544doi: 10.1038/nprot.2006.468google scholar: lookup
  23. Gundry RL, White MY, Murray CI, Kane LA, Fu Q, Stanley BA, Van Eyk JE. Preparation of proteins and peptides for mass spectrometry analysis in a bottom-up proteomics workflow.. Curr Protoc Mol Biol 2009 Oct;Chapter 10:Unit10.25.
    doi: 10.1002/0471142727.mb1025s88pmc: PMC2905857pubmed: 19816929google scholar: lookup
  24. Cox J, Mann M. MaxQuant enables high peptide identification rates, individualized p.p.b.-range mass accuracies and proteome-wide protein quantification.. Nat Biotechnol 2008 Dec;26(12):1367-72.
    pubmed: 19029910doi: 10.1038/nbt.1511google scholar: lookup
  25. Cox J, Neuhauser N, Michalski A, Scheltema RA, Olsen JV, Mann M. Andromeda: a peptide search engine integrated into the MaxQuant environment.. J Proteome Res 2011 Apr 1;10(4):1794-805.
    pubmed: 21254760doi: 10.1021/pr101065jgoogle scholar: lookup
  26. Tyanova S, Temu T, Cox J. The MaxQuant computational platform for mass spectrometry-based shotgun proteomics.. Nat Protoc 2016 Dec;11(12):2301-2319.
    pubmed: 27809316doi: 10.1038/nprot.2016.136google scholar: lookup
  27. Hashimoto M, Nambo Y, Kondo T, Watanabe K, Orino K. A Study on the Presence of Ferritin-binding Proteins in Fetal Horse Plasma.. J Equine Sci 2011;22(1):1-7.
    pmc: PMC4013997pubmed: 24833981doi: 10.1294/jes.22.1google scholar: lookup
  28. Needleman SB, Wunsch CD. A general method applicable to the search for similarities in the amino acid sequence of two proteins.. J Mol Biol 1970 Mar;48(3):443-53.
    pubmed: 5420325doi: 10.1016/0022-2836(70)90057-4google scholar: lookup
  29. nMadden T. The BLAST sequence analysis tool. The NCBI Handbook 2nd ed.nBethesda, MD: National Centre for Biotechnology Information (US); 2013.
  30. Altschul SF, Madden TL, Schäffer AA, Zhang J, Zhang Z, Miller W, Lipman DJ. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs.. Nucleic Acids Res 1997 Sep 1;25(17):3389-402.
    pmc: PMC146917pubmed: 9254694doi: 10.1093/nar/25.17.3389google scholar: lookup
  31. Liu H, Sadygov RG, Yates JR 3rd. A model for random sampling and estimation of relative protein abundance in shotgun proteomics.. Anal Chem 2004 Jul 15;76(14):4193-201.
    pubmed: 15253663doi: 10.1021/ac0498563google scholar: lookup

Citations

This article has been cited 1 times.
  1. Brown JE, Noormohammadi AH, Courtman NF. Immunoreactivity of canine, feline, and equine D-dimer with antibodies to human D-dimer. J Vet Intern Med 2024 Jan-Feb;38(1):187-196.
    doi: 10.1111/jvim.16888pubmed: 37950415google scholar: lookup
Subscribe to our newsletter
Join 100,127 horse owners like you who receive our email updates on horse health and nutrition.